Advancements in immunosuppressive drugs, surgical techniques, and postoperative care have significantly improved outcomes of liver transplantation;however, cold preservation-associated ischemia/reperfusion (I/R) injury remains a major problem complicating posttransplant care and subsequent long-term transplant outcomes. Further, the recent organ shortage demands to expand the donor pool, and the use of liver grafts from marginal donors possibly augments I/R injury. Carbon monoxide (CO), an endogenous byproduct of heme, has lately received notable attention as a regulatory molecule in cellular and biological processes. During the last 3 years, we have evaluated the cytoprotective function of CO and obtained encouraging results showing that exogenous inhaled CO provides benefits against transplant-induced hepatic I/R injury. We hypothesize that CO protects liver grafts from hepatic I/R injury via the regulation of proinflammatory responses by directly inhibiting Kupffer cell activation and by improving sinusoidal circulation. We will pursue two specific aims to explore the clinical applicability, efficacy, and mechanisms of protection of low dose inhaled CO in ameliorating liver damage due to transplant preservation injury. AIM I: TO OPTIMIZE INHALED CO DELIVERY TO INHIBIT LIVER I/R INJURY. We will fully explore the regulatory effects of inhaled CO delivered in a clinically applicable manner to ameliorate hepatic I/R injury. Optimization of the delivery protocol will be established by administering CO as a brief inhalation therapy during the peritransplant period. To maximize CO efficacy, we will study effects obtained with donor and/or recipient treatment with CO. The potential adverse consequences of CO inhalation will also be investigated. AIM II: TO IDENTIFY THE MECHANISMS OF CO-MEDIATED PROTECTION OF LIVER GRAFTS. We will determine Kupffer cells as CO's direct target for antiproinflammatory function. Molecular mechanisms of CO- mediated antiinflammatory protection via the MAPK signaling pathways will be analyzed in isolated cell population. We will determine the mechanism of CO-mediated SEC protection and hepatic sinusoidal circulation via the inhibition of stellate cell activation and endothelin (ET) system. Our long-term goal in this proposal is to harness the benefit of CO pathway to improve early liver graft function and minimize the morbidity associated with severe preservation injury by fully exploring the regulatory mechanisms of CO.